Inotropic response to β-adrenergic receptor stimulation and anti-adrenergic effect of ACh in endothelial NO synthase-deficient mouse hearts

1. The functional consequences of a lack of endothelial nitric oxide synthase (eNOS) on left ventricular Force development and the anti-adrenergic gic effect of acetylcholine (ACh) M ere investigated in isolated hearts and cardiomyocytes from wild type (WT) and eNOS knockout (eNOS-/-) mice. 2. eNOS expression in cardiac myocytes accounted for 20% of total cardiac eNOS (Western blot analysis). These results were confirmed hv RT-PCR analysis. 3. In the unstimulated perfused heart, the left ventricular pressure (LVP) and maximal rate of left ventricular force development (dP/dt(max)) of eNOS-/- hearts were not significantly different from those of WT hearts (LVP: 97 +/- 11 mmHg WT vs. 111 +/- 11 mmHg eNOS-/-; dP/dt(max): 3700 +/- 712 mmHg s(-1) WT vs. 4493 +/- 320 mmHg s(-1) eNOS-/-). 4. The dobutamine (10-300 nM)-induced increase in LVP was enhanced in eNOS-/- hearts. In contrast, L-tvpe Ca2+ currents (I-Ca,I-L) in isolated cardiomyocytes of WT and eNOS-/- heal ts showed no differences after beta -adrenergic stimulation. Dibutyryl-cGMP (50 muM) reduced basal I-Ca,I-L in WT cells to 72 +/- 12% while eNOS-/- I-Ca,I-L was insensitive to the drug. The pre stimulated I-Ca,I-L (30 nM isoproterenol) was attenuated by dibutyryl-cGMP in WT and eNOS-/- cells to the same extent. 5. The Ca2+ (1.5-4.5 mM)-induced increase in inotropy was not different between the two experimental groups and P-adrenergic receptor density was increased by 50% in eNOS-/- hearts. 6. The contractile effects of dobutamine could be inhibited almost completely by ACh ol adenosine. The extent of the anti-adrenergic effect of both compounds was identical in WT and eNOS-/- hearts. Measurement of I-Ca,I-L in isolated cardiac myocytes yielded similar results. 7. These data demonstrate that in the Adult mouse (1) lack of eNOS is associated with increased cardiac contractile force in response to beta -adrenergic stimulation and with elevated beta -adrenergic receptor density, (2) the unaltered response of I-Ca,I-L in eNOS-/- cardiac myocytes to beta -adrenergic stimulation suggests that endothelium-derived NO is important in mediating the whole-organ effects and (3) eNOS is unimportant fur the anti-adrenergic effect of ACh and adenosine.